ResearchPad - regulatory-proteins https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[A MYB transcription factor, <i>BnMYB2</i>, cloned from ramie (<i>Boehmeria nivea</i>) is involved in cadmium tolerance and accumulation]]> https://www.researchpad.co/article/elastic_article_15762 MYB-related transcription factors play important roles in plant development and response to various environmental stresses. In the present study, a novel MYB gene, designated as BnMYB2 (GenBank accession number: MF741319.1), was isolated from Boehmeria nivea using rapid amplification of cDNA ends (RACE) and RT-PCR on a sequence fragment from a ramie transcriptome. BnMYB2 has a 945 bp open reading frame encoding a 314 amino acid protein that contains a DNA-binding domain and shares high sequence identity with MYB proteins from other plant species. The BnMYB2 promoter contains several putative cis-acting elements involved in stress or phytohormone responses. A translational fusion of BnMYB2 with enhanced green fluorescent protein (eGFP) showed nuclear and cytosolic subcellular localization. Real-time PCR results indicated that BnMYB2 expression was induced by Cadmium (Cd) stress. Overexpression of BnMYB2 in Arabidopsis thaliana resulted in a significant increase of Cd tolerance and accumulation. Thus, BnMYB2 positively regulated Cd tolerance and accumulation in Arabidopsis, and could be used to enhance the efficiency of Cd removal with plants.

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<![CDATA[Single-nucleus RNA-seq identifies divergent populations of FSHD2 myotube nuclei]]> https://www.researchpad.co/article/elastic_article_14649 Although misexpression of DUX4 has been known as the major cause in FSHD, it is lowly expressed in patient samples and analysis of the consequences of DUX4 expression has largely relied on artificial overexpression. Here, we took advantage of recent methodological advances to observe native DUX4 expression at the single-nucleus level in FSHD2 patient-derived myotubes. Using single-nucleus RNA-seq (snRNA-seq), we were able to detect endogenous DUX4-expressing nuclei and the extent of spreading of DUX4-target gene expression across many nuclei. Our highly sensitive snRNA-seq method further allowed us to identify two populations of FSHD myotube nuclei with distinct transcriptional profiles. One is highly enriched with DUX4 and target genes (FSHD-Hi) while the other has sparser DUX4 and FSHD-induced genes expressed (FSHD-Lo), reflecting two potentially different pathological states of patient myotubes. We observed a set of transcription factors specifically upregulated in FSHD-Hi myotube nuclei associated with the cell cycle, and significant upregulation of DUX4 paralog DUXA that contributes to further upregulation of DUX4 target genes. We propose that transcription factors downstream of DUX4 may amplify DUX4 signal and thus act to perpetuate FSHD.

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<![CDATA[Identification of a new R3 MYB type repressor and functional characterization of the members of the MBW transcriptional complex involved in anthocyanin biosynthesis in eggplant (<i>S</i>. <i>melongena</i> L.)]]> https://www.researchpad.co/article/elastic_article_14583 Here we focus on the highly conserved MYB-bHLH-WD repeat (MBW) transcriptional complex model in eggplant, which is pivotal in the transcriptional regulation of the anthocyanin biosynthetic pathway. Through a genome-wide approach performed on the recently released Eggplant Genome (cv. 67/3) previously identified, and reconfirmed by us, members belonging to the MBW complex (SmelANT1, SmelAN2, SmelJAF13, SmelAN1) were functionally characterized. Furthermore, a regulatory R3 MYB type repressor (SmelMYBL1), never reported before, was identified and characterized as well.

Through a qPCR approach, we revealed specific transcriptional patterns of candidate genes in different plant tissue/organs at two stages of fruit development. Two strategies were adopted for investigating the interactions of bHLH partners (SmelAN1, SmelJAF13) with MYB counterparts (SmelANT1, SmelAN2 and SmelMYBL1): Yeast Two Hybrid (Y2H) and Bimolecular Fluorescent Complementation (BiFC) in A. thaliana mesophylls protoplast. Agro-infiltration experiments highlighted that N. benthamiana leaves transiently expressing SmelANT1 and SmelAN2 showed an anthocyanin-pigmented phenotype, while their co-expression with SmelMYBL1 prevented anthocyanin accumulation. Our results suggest that SmelMYBL1 may inhibits the MBW complex via the competition with MYB activators for bHLH binding site, although this hypothesis requires further elucidation.

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<![CDATA[HSPA6 augments garlic extract-induced inhibition of proliferation, migration, and invasion of bladder cancer EJ cells; Implication for cell cycle dysregulation, signaling pathway alteration, and transcription factor-associated MMP-9 regulation]]> https://www.researchpad.co/article/5989db4fab0ee8fa60bdb9fe

Although recent studies have demonstrated the anti-tumor effects of garlic extract (GE), the exact molecular mechanism is still unclear. In this study, we investigated the molecular mechanism associated with the inhibitory action of GE against bladder cancer EJ cell responses. Treatment with GE significantly inhibited proliferation of EJ cells dose-dependently through G2/M-phase cell cycle arrest. This G2/M-phase cell cycle arrest by GE was due to the activation of ATM and CHK2, which appears to inhibit phosphorylation of Cdc25C (Ser216) and Cdc2 (Thr14/Tyr15), this in turn was accompanied by down-regulation of cyclin B1 and up-regulation of p21WAF1. Furthermore, GE treatment was also found to induce phosphorylation of MAPK (ERK1/2, p38MAPK, and JNK) and AKT. In addition, GE impeded the migration and invasion of EJ cells via inhibition of MMP-9 expression followed by decreased binding activities of AP-1, Sp-1, and NF-κB motifs. Based on microarray datasets, we selected Heat shock protein A6 (HSPA6) as the most up-regulated gene responsible for the inhibitory effects of GE. Interestingly, overexpression of HSPA6 gene resulted in an augmentation effect with GE inhibiting proliferation, migration, and invasion of EJ cells. The augmentation effect of HSPA6 was verified by enhancing the induction of G2/M-phase-mediated ATM-CHK2-Cdc25C-p21WAF1-Cdc2 cascade, phosphorylation of MAPK and AKT signaling, and suppression of transcription factor-associated MMP-9 regulation in response to GE in EJ cells. Overall, our novel results indicate that HSPA6 reinforces the GE-mediated inhibitory effects of proliferation, migration, and invasion of EJ cells and may provide a new approach for therapeutic treatment of malignancies.

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<![CDATA[Podocyte RNA sequencing reveals Wnt- and ECM-associated genes as central in FSGS]]> https://www.researchpad.co/article/Nff231b2e-f2d8-47eb-acf2-c510faf35a1a

Loss of podocyte differentiation can cause nephrotic-range proteinuria and Focal and Segmental Glomerulosclerosis (FSGS). As specific therapy is still lacking, FSGS frequently progresses to end-stage renal disease. The exact molecular mechanisms of FSGS and gene expression changes in podocytes are complex and widely unknown as marker changes have mostly been assessed on the glomerular level. To gain a better insight, we isolated podocytes of miR-193a overexpressing mice, which suffer from FSGS due to suppression of the podocyte master regulator Wt1. We characterised the podocytic gene expression changes by RNAseq and identified many novel candidate genes not linked to FSGS so far. This included strong upregulation of the receptor tyrosine kinase EphA6 and a massive dysregulation of circadian genes including the loss of the transcriptional activator Arntl. By comparison with podocyte-specific changes in other FSGS models we found a shared dysregulation of genes associated with the Wnt signaling cascade, while classical podocyte-specific genes appeared widely unaltered. An overlap with gene expression screens from human FSGS patients revealed a strong enrichment in genes associated with extra-cellular matrix (ECM) and metabolism. Our data suggest that FSGS progression might frequently depend on pathways that are often overlooked when considering podocyte homeostasis.

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<![CDATA[Unusual genome expansion and transcription suppression in ectomycorrhizal Tricholoma matsutake by insertions of transposable elements]]> https://www.researchpad.co/article/Nd7412b83-0508-48a9-959e-b3aa8ede7a25

Genome sequencing of Tricholoma matsutake revealed its unusually large size as 189.0 Mbp, which is a consequence of extraordinarily high transposable element (TE) content. We identified that 702 genes were surrounded by TEs, and 83.2% of these genes were not transcribed at any developmental stage. This observation indicated that the insertion of TEs alters the transcription of the genes neighboring these TEs. Repeat-induced point mutation, such as C to T hypermutation with a bias over “CpG” dinucleotides, was also recognized in this genome, representing a typical defense mechanism against TEs during evolution. Many transcription factor genes were activated in both the primordia and fruiting body stages, which indicates that many regulatory processes are shared during the developmental stages. Small secreted protein genes (<300 aa) were dominantly transcribed in the hyphae, where symbiotic interactions occur with the hosts. Comparative analysis with 37 Agaricomycetes genomes revealed that IstB-like domains (PF01695) were conserved across taxonomically diverse mycorrhizal genomes, where the T. matsutake genome contained four copies of this domain. Three of the IstB-like genes were overexpressed in the hyphae. Similar to other ectomycorrhizal genomes, the CAZyme gene set was reduced in T. matsutake, including losses in the glycoside hydrolase genes. The T. matsutake genome sequence provides insight into the causes and consequences of genome size inflation.

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<![CDATA[Autosomal recessive congenital cataracts linked to HSF4 in a consanguineous Pakistani family]]> https://www.researchpad.co/article/Na302ecef-6336-4a97-9663-2461453833de

Purpose

To investigate the genetic basis of autosomal recessive congenital cataracts (arCC) in a large consanguineous Pakistani family.

Methods

All participating members of family, PKCC074 underwent an ophthalmic examination. Slit-lamp photographs were ascertained for affected individuals that have not been operated for the removal of the cataractous lens. A small aliquot of the blood sample was collected from all participating individuals and genomic DNAs were extracted. A genome-wide scan was performed with polymorphic short tandem repeat (STR) markers and the logarithm of odds (LOD) scores were calculated. All coding exons and exon-intron boundaries of HSF4 were sequenced and expression of Hsf4 in mouse ocular lens was investigated. The C-terminal FLAG-tagged wild-type and mutant HSF4b constructs were prepared to examine the nuclear localization pattern of the mutant protein.

Results

The ophthalmological examinations suggested that nuclear cataracts are present in affected individuals. Genome-wide linkage analyses localized the critical interval to a 10.95 cM (14.17 Mb) interval on chromosome 16q with a maximum two-point LOD score of 4.51 at θ = 0. Sanger sequencing identified a novel missense mutation: c.433G>C (p.Ala145Pro) that segregated with the disease phenotype in the family and was not present in ethnically matched controls. Real-time PCR analysis identified the expression of HSF4 in mouse lens as early as embryonic day 15 with a steady level of expression thereafter. The immunofluorescence tracking confirmed that both wild-type and mutant HSF4 (p.Ala145Pro) proteins localized to the nucleus.

Conclusion

Here, we report a novel missense mutation in HSF4 associated with arCC in a familial case of Pakistani descent.

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<![CDATA[Identification and expression profiling of miRNAs in two color variants of carrot (Daucus carota L.) using deep sequencing]]> https://www.researchpad.co/article/5c8accd2d5eed0c48499009d

microRNAs represent small endogenous RNAs which are known to play a crucial role in various plant metabolic processes. Carrot being an important vegetable crop, represents one of the richest sources of carotenoids and anthocyanins. Most of the studies on microRNAs have been conducted in the aerial parts of the plants. However, carrot has the rare distinction of storing these compounds in roots. Therefore, carrot represents a good model system to unveil the regulatory roles of miRNAs in the underground edible part of the plant. For the first time, we report the genome wide identification and expression profiling of miRNAs in two contrasting color variants of carrot namely Orange Red and Purple Black using RNA-seq. Illumina sequencing resulted in the generation of 25.5M and 18.9M reads in Orange Red and Purple Black libraries, respectively. In total, 144 and 98 (read count >10), conserved microRNAs and 36 and 66 novel microRNAs were identified in Orange Red and Purple Black, respectively. Functional categorization and differential gene expression revealed the presence of several miRNA genes targeting various secondary metabolic pathways including carotenoid and anthocyanin biosynthetic pathways in the two libraries. 11 known and 2 novel microRNAs were further validated using Stem-Loop PCR and qRT-PCR. Also, target validation was performed for selected miRNA genes using RLM-RACE approach. The present work has laid a foundation towards understanding of various metabolic processes, particularly the color development in carrot. This information can be further employed in targeted gene expression for increasing the carotenoid and anthocyanin content in crop plants.

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<![CDATA[A paper-based, cell-free biosensor system for the detection of heavy metals and date rape drugs]]> https://www.researchpad.co/article/5c897715d5eed0c4847d23fa

Biosensors have emerged as a valuable tool with high specificity and sensitivity for fast and reliable detection of hazardous substances in drinking water. Numerous substances have been addressed using synthetic biology approaches. However, many proposed biosensors are based on living, genetically modified organisms and are therefore limited in shelf life, usability and biosafety. We addressed these issues by the construction of an extensible, cell-free biosensor. Storage is possible through freeze drying on paper. Following the addition of an aqueous sample, a highly efficient cell-free protein synthesis (CFPS) reaction is initiated. Specific allosteric transcription factors modulate the expression of ‘superfolder’ green fluorescent protein (sfGFP) depending on the presence of the substance of interest. The resulting fluorescence intensities are analyzed with a conventional smartphone accompanied by simple and cheap light filters. An ordinary differential equitation (ODE) model of the biosensors was developed, which enabled prediction and optimization of performance. With an optimized cell-free biosensor based on the Shigella flexneri MerR transcriptional activator, detection of 6 μg/L Hg(II) ions in water was achieved. Furthermore, a completely new biosensor for the detection of gamma-hydroxybutyrate (GHB), a substance used as date-rape drug, was established by employing the naturally occurring transcriptional repressor BlcR from Agrobacterium tumefaciens.

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<![CDATA[O-GlcNAcylation of PERIOD regulates its interaction with CLOCK and timing of circadian transcriptional repression]]> https://www.researchpad.co/article/5c5ca281d5eed0c48441e509

Circadian clocks coordinate time-of-day-specific metabolic and physiological processes to maximize organismal performance and fitness. In addition to light and temperature, which are regarded as strong zeitgebers for circadian clock entrainment, metabolic input has now emerged as an important signal for clock entrainment and modulation. Circadian clock proteins have been identified to be substrates of O-GlcNAcylation, a nutrient sensitive post-translational modification (PTM), and the interplay between clock protein O-GlcNAcylation and other PTMs is now recognized as an important mechanism by which metabolic input regulates circadian physiology. To better understand the role of O-GlcNAcylation in modulating clock protein function within the molecular oscillator, we used mass spectrometry proteomics to identify O-GlcNAcylation sites of PERIOD (PER), a repressor of the circadian transcriptome and a critical biochemical timer of the Drosophila clock. In vivo functional characterization of PER O-GlcNAcylation sites indicates that O-GlcNAcylation at PER(S942) reduces interactions between PER and CLOCK (CLK), the key transcriptional activator of clock-controlled genes. Since we observe a correlation between clock-controlled daytime feeding activity and higher level of PER O-GlcNAcylation, we propose that PER(S942) O-GlcNAcylation during the day functions to prevent premature initiation of circadian repression phase. This is consistent with the period-shortening behavioral phenotype of per(S942A) flies. Taken together, our results support that clock-controlled feeding activity provides metabolic signals to reinforce light entrainment to regulate circadian physiology at the post-translational level. The interplay between O-GlcNAcylation and other PTMs to regulate circadian physiology is expected to be complex and extensive, and reach far beyond the molecular oscillator.

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<![CDATA[Mapping DNA sequence to transcription factor binding energy in vivo]]> https://www.researchpad.co/article/5c61e8e5d5eed0c48496f361

Despite the central importance of transcriptional regulation in biology, it has proven difficult to determine the regulatory mechanisms of individual genes, let alone entire gene networks. It is particularly difficult to decipher the biophysical mechanisms of transcriptional regulation in living cells and determine the energetic properties of binding sites for transcription factors and RNA polymerase. In this work, we present a strategy for dissecting transcriptional regulatory sequences using in vivo methods (massively parallel reporter assays) to formulate quantitative models that map a transcription factor binding site’s DNA sequence to transcription factor-DNA binding energy. We use these models to predict the binding energies of transcription factor binding sites to within 1 kBT of their measured values. We further explore how such a sequence-energy mapping relates to the mechanisms of trancriptional regulation in various promoter contexts. Specifically, we show that our models can be used to design specific induction responses, analyze the effects of amino acid mutations on DNA sequence preference, and determine how regulatory context affects a transcription factor’s sequence specificity.

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<![CDATA[Transvection-like interchromosomal interaction is not observed at the transcriptional level when tested in the Rosa26 locus in mouse]]> https://www.researchpad.co/article/5c6f1494d5eed0c48467a353

Long-range associations between enhancers and their target gene promoters have been shown to play critical roles in executing genome function. Recent variations of chromosome capture technology have revealed a comprehensive view of intra- and interchromosomal contacts between specific genomic sites. The locus control region of the β-globin genes (β-LCR) is a super-enhancer that is capable of activating all of the β-like globin genes within the locus in cis through physical interaction by forming DNA loops. CTCF helps to mediate loop formation between LCR-HS5 and 3’HS1 in the human β-globin locus, in this way thought to contribute to the formation of a “chromatin hub”. The β-globin locus is also in close physical proximity to other erythrocyte-specific genes located long distances away on the same chromosome. In this case, erythrocyte-specific genes gather together at a shared “transcription factory” for co-transcription. Theoretically, enhancers could also activate target gene promoters at the identical loci, yet on different chromosomes in trans, a phenomenon originally described as transvection in Drosophilla. Although close physical proximity has been reported for the β-LCR and the β-like globin genes when integrated at the mouse homologous loci in trans, their structural and functional interactions were found to be rare, possibly because of a lack of suitable regulatory elements that might facilitate such trans interactions. Therefore, we re-evaluated presumptive transvection-like enhancer-promoter communication by introducing CTCF binding sites and erythrocyte-specific transcription units into both LCR-enhancer and β-promoter alleles, each inserted into the mouse ROSA26 locus on separate chromosomes. Following cross-mating of mice to place the two mutant loci at the identical chromosomal position and into active chromation in trans, their transcriptional output was evaluated. The results demonstrate that there was no significant functional association between the LCR and the β-globin gene in trans even in this idealized experimental context.

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<![CDATA[Context dependent roles for RB-E2F transcriptional regulation in tumor suppression]]> https://www.researchpad.co/article/5c5ca2b7d5eed0c48441e94f

RB-E2F transcriptional control plays a key role in regulating the timing of cell cycle progression from G1 to S-phase in response to growth factor stimulation. Despite this role, it is genetically dispensable for cell cycle exit in primary fibroblasts in response to growth arrest signals. Mice engineered to be defective for RB-E2F transcriptional control at cell cycle genes were also found to live a full lifespan with no susceptibility to cancer. Based on this background we sought to probe the vulnerabilities of RB-E2F transcriptional control defects found in Rb1R461E,K542E mutant mice (Rb1G) through genetic crosses with other mouse strains. We generated Rb1G/G mice in combination with Trp53 and Cdkn1a deficiencies, as well as in combination with KrasG12D. The Rb1G mutation enhanced Trp53 cancer susceptibility, but had no effect in combination with Cdkn1a deficiency or KrasG12D. Collectively, this study indicates that compromised RB-E2F transcriptional control is not uniformly cancer enabling, but rather has potent oncogenic effects when combined with specific vulnerabilities.

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<![CDATA[Identification of new regulators through transcriptome analysis that regulate anthocyanin biosynthesis in apple leaves at low temperatures]]> https://www.researchpad.co/article/5c59fef2d5eed0c4841357fd

Anthocyanin pigments play many roles in plants, including providing protection against biotic and abiotic stresses. To identify new regulatory genes in apple (Malus domestica) that may be involved in regulating low temperature induced anthocyanin biosynthesis, we performed RNA-seq analysis of leaves from the ‘Gala’ apple cultivar following exposure to a low temperature (16 °C). A visible red color appeared on the upper leaves and the anthocyanin content increased significantly after the low temperature treatment. Genes from the flavonoid biosynthesis pathway were significantly enriched among the differentially expressed genes, and the expression of several transcription factors was shown by WGCNA (weighted gene co-expression network analysis) to correlate with anthocyanin accumulation, including members of the MYB, MADS, WRKY, WD40, Zinc Finger and HB-ZIP families. Three MYB transcription factors (MdMYB12, MdMYB22 and MdMYB114), which had several CBF/DREB response elements in their promoters, were significantly induced by low temperature exposure and their expression also correlated highly with anthocyanin accumulation. We hypothesize that they may act as regulators of anthocyanin biosynthesis and be regulated by CBF/DREB transcription factors in apple leaves under low temperature conditions. The analyses presented here provide insights into the molecular mechanisms underlying anthocyanin accumulation during low temperature exposure.

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<![CDATA[The inhibitor apoptosis protein antagonist Debio 1143 Is an attractive HIV-1 latency reversal candidate]]> https://www.researchpad.co/article/5c61e8bbd5eed0c48496f07d

Antiretroviral therapy (ART) suppresses HIV replication, but does not cure the infection because replication-competent virus persists within latently infected CD4+ T cells throughout years of therapy. These reservoirs contain integrated HIV-1 genomes and can resupply active virus. Thus, the development of strategies to eliminate the reservoir of latently infected cells is a research priority of global significance. In this study, we tested efficacy of a new inhibitor of apoptosis protein antagonist (IAPa) called Debio 1143 at reversing HIV latency and investigated its mechanisms of action. Debio 1143 activates HIV transcription via NF-kB signaling by degrading the ubiquitin ligase baculoviral IAP repeat-containing 2 (BIRC2), a repressor of the non-canonical NF-kB pathway. Debio 1143-induced BIRC2 degradation results in the accumulation of NF-κB-inducing kinase (NIK) and proteolytic cleavage of p100 into p52, leading to nuclear translocation of p52 and RELB. Debio 1143 greatly enhances the binding of RELB to the HIV-1 LTR. These data indicate that Debio 1143 activates the non-canonical NF-kB signaling pathway by promoting the binding of RELB:p52 complexes to the HIV-1 LTR, resulting in the activation of the LTR-dependent HIV-1 transcription. Importantly, Debio 1143 reverses viral latency in HIV-1 latent T cell lines. Using knockdown (siRNA BIRC2), knockout (CRIPSR NIK) and proteasome machinery neutralization (MG132) approaches, we found that Debio 1143-mediated HIV latency reversal is BIRC2 degradation- and NIK stabilization-dependent. Debio 1143 also reverses HIV-1 latency in resting CD4+ T cells derived from ART-treated patients or HIV-1-infected humanized mice under ART. Interestingly, daily oral administration of Debio 1143 in cancer patients at well-tolerated doses elicited BIRC2 target engagement in PBMCs and induced a moderate increase in cytokines and chemokines mechanistically related to NF-kB signaling. In conclusion, we provide strong evidences that the IAPa Debio 1143, by initially activating the non-canonical NF-kB signaling and subsequently reactivating HIV-1 transcription, represents a new attractive viral latency reversal agent (LRA).

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<![CDATA[Finding the needle in a haystack: Mapping antifungal drug resistance in fungal pathogen by genomic approaches]]> https://www.researchpad.co/article/5c5ca2fad5eed0c48441eee3 ]]> <![CDATA[Epicatechin modulates stress-resistance in C. elegans via insulin/IGF-1 signaling pathway]]> https://www.researchpad.co/article/5c58d64cd5eed0c484031b1e

The nematode Caenorhabditis elegans has been used to examine the influence of epicatechin (EC), an abundant flavonoid in the human diet, in some stress biomarkers (ROS production, lipid peroxidation and protein carbonylation). Furthermore, the ability of EC to modulate the expression of some key genes in the insulin/IGF-1 signaling pathway (IIS), involved in longevity and oxidative or heat shock stress response, has also been explored. The final aim was to contribute to the elucidation of the mechanisms involved in the biological effects of flavonoids. The results showed that EC-treated wild-type C. elegans exhibited increased survival and reduced oxidative damage of biomolecules when submitted to thermal stress. EC treatment led to a moderate elevation in ROS levels, which might activate endogenous mechanisms of defense protecting against oxidative insult. The enhanced stress resistance induced by EC was found to be mediated through the IIS pathway, since assays in daf-2, age-1, akt-1, akt-2, sgk-1, daf-16, skn-1 and hsf-1 loss of function mutant strains failed to show any heat-resistant phenotype against thermal stress when treated with EC. Consistently, EC treatment upregulated the expression of some stress resistance associated genes, such as gst-4, hsp-16.2 and hsp-70, which are downstream regulated by the IIS pathway.

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<![CDATA[Unique and overlapping GLI1 and GLI2 transcriptional targets in neoplastic chondrocytes]]> https://www.researchpad.co/article/5c59ff07d5eed0c48413599d

Excessive Hedgehog (Hh) signaling in chondrocytes is sufficient to cause formation of enchondroma-like lesions which can progress to chondrosarcoma. To elucidate potential underlying mechanisms, we identified GLI1 and GLI2 target genes in human chondrosarcoma. Using chromatin immunoprecipitation (ChIP) sequencing and microarray data, in silico analyses were conducted to identify and characterize unique and overlapping GLI1 and GLI2 binding regions in neoplastic chondrocytes. After overlaying microarray data from human chondrosarcoma, 204 upregulated and 106 downregulated genes were identified as Hh-responsive Gli binding targets. After overlaying published Gli ChIP-on-chip data from mouse, 48 genes were identified as potential direct downstream targets of Hedgehog signaling with shared GLI binding regions in evolutionarily conserved DNA elements. Among these was BMP2, pointing to potential cross-talk between TGF beta signaling and Hh signaling. Our identification of potential target genes that are unique and common to GLI1 and GLI2 in neoplastic chondrocytes contributes to elucidating potential pathways through which Hh signaling impacts cartilage tumor biology.

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<![CDATA[Multi-study inference of regulatory networks for more accurate models of gene regulation]]> https://www.researchpad.co/article/5c536a85d5eed0c484a47592

Gene regulatory networks are composed of sub-networks that are often shared across biological processes, cell-types, and organisms. Leveraging multiple sources of information, such as publicly available gene expression datasets, could therefore be helpful when learning a network of interest. Integrating data across different studies, however, raises numerous technical concerns. Hence, a common approach in network inference, and broadly in genomics research, is to separately learn models from each dataset and combine the results. Individual models, however, often suffer from under-sampling, poor generalization and limited network recovery. In this study, we explore previous integration strategies, such as batch-correction and model ensembles, and introduce a new multitask learning approach for joint network inference across several datasets. Our method initially estimates the activities of transcription factors, and subsequently, infers the relevant network topology. As regulatory interactions are context-dependent, we estimate model coefficients as a combination of both dataset-specific and conserved components. In addition, adaptive penalties may be used to favor models that include interactions derived from multiple sources of prior knowledge including orthogonal genomics experiments. We evaluate generalization and network recovery using examples from Bacillus subtilis and Saccharomyces cerevisiae, and show that sharing information across models improves network reconstruction. Finally, we demonstrate robustness to both false positives in the prior information and heterogeneity among datasets.

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<![CDATA[Physical stimulation by REAC and BMP4/WNT-1 inhibitor synergistically enhance cardiogenic commitment in iPSCs]]> https://www.researchpad.co/article/5c5217b6d5eed0c48479441d

It is currently known that pluripotent stem cells can be committed in vitro to the cardiac lineage by the modulation of specific signaling pathways, but it is also well known that, despite the significant increase in cardiomyocyte yield provided by the currently available conditioned media, the resulting cardiogenic commitment remains a highly variable process. Previous studies provided evidence that radio electric fields asymmetrically conveyed through the Radio Electric Asymmetric Conveyer (REAC) technology are able to commit R1 embryonic stem cells and human adipose derived stem cells toward a cardiac phenotype. The present study aimed at investigating whether the effect of physical stimulation by REAC in combination with specific chemical inductors enhance the cardiogenic potential in human induced pluripotent stem cells (iPSCs). The appearance of a cardiac-like phenotype in iPSCs cultured in the presence of a cardiogenic medium, based upon BMP4 and a WNT-inhibitor, was consistently increased by REAC treatment used only during the early fate differentiation for the first 72 hours. REAC-exposed iPSCs exhibited an upregulation in the expression of specific cardiogenic transcripts and morphologically in the number of beating clusters, as compared to cells cultured in the cardiogenic medium alone. Our results indicate that physical modulation of cellular dynamics provided by the REAC offers an affordable strategy to mimic iPSC cardiac-like fates in the presence of a cardiogenic milieu.

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